A cell surface glycoprotein, fibronectin, is involved in cell adhesion in human skin fibroblasts. Fibronectin is not involved in the initiation of cell proliferation. During cellular aging changes at the cell surface and in the fibronectin have been reported. This proposal will evaluate by labelling and immunoprecipitation, lactoperoxidase catalyzed iodination and immunofluorescence the amount of intracellular, cell surface associated and secreted fibronectin in human skin fibroblasts before and after the cells lose their proliferative capacity. In order to determine the events which regulate the fibronectin in human skin fibroblasts during in vitro aging the kinetics of synthesis, transit to the cell surface, turnover and secretion will be determined. Further information on the control of synthesis of fibronectin will be obtained by measuring the levels of translatable mRNA for fibronectin in a cell free system. The amounts and specific activity of cellular proteases in aging human skin fibroblasts will increase our understanding of the degradation process. The production of large amounts of fibronectin also provides a unique opportunity to evaluate the structure of one of the major glycoproteins of the extracellular matrix in human skin fibroblasts during in vitro cellular aging. Peptides profiles of fibronectin peptides produced by proteolysis will be compared before and after the cells have lost their proliferative capacity. Posttranslational modification of fibronectin during celluar aging will also be evaluated. This proposal will investigate the regulation and structure of one of the major glycoproteins of human skin fibroblasts during cellular aging. The information provided in this proposal may be essential towards our understanding of the mechanism reulting in the loss of proliferative capacity of normal human fibroblasts-like cells in vitro and in vivo.